Anthrax is a high risk bioterrorist threat. Despite high dose antibiotic therapy, several deaths occurred following the 2001 mail attack. Passive immunization with anti-anthrax immunoglobulins, in combination with antibiotics, is a promising approach for post-exposure treatment. Hematech is developing a genetically modified line of cattle that will produce human instead of bovine immunoglobulin. These cattle may be hyperimmunized with anthrax antigens and produce highly efficacious polyclonal antibodies for protection against anthrax, and many other bioterrorist agents. In phase I, we introduced a human artificial chromosome containing the entire sequences of human immunoglobulin heavy and light chain genes into bovine fibroblasts and produced transchromosomic (Tc) cattle using somatic cell cloning technology. We showed that Tc calves retain the HAC and produce low levels of human IgG in blood (2-30 mg/L). We also developed an immunization scheme, in wild wildtype Holstein steers, using various antigen-adjuvant formulations and booster vaccinations to produce high titer antibodies to anthrax. Furthermore, we developed and optimized ELISA assays, which were used to detect IgG titers to anthrax protective antigen, edema factor and lethal factor antigens, and we refined an in-vitro toxin neutralization assay, to quantify and demonstrate the biological potency of the antibodies. Under Phase II support we will vaccinate and boost Tc cattle produced in Phase I, collect plasma and purify human and bovine antibodies separately. We will refine an in vivo mouse protection assay and evaluate the therapeutic potency of the purified human antibody as compared to purified bovine antibody. We will also produce Tc calves in which the bovine heavy chain genes have been knocked out in an effort to increase levels of human immunoglobulin production. The ultimate goal of phase II is to complete the development of a bovine system for production of human polyclonal antibodies with efficacy in the treatment of anthrax infections.